Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.4 (trypsin)
42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

This paper presents morphological data on mouse oocyte maturation and fertilization, reviews evidence supporting the existence of a sperm receptor, and suggests future directions for this line of research. We used scanning electron microscopy to examine oocytes under a variety of conditions. The surfaces of mature mouse oocytes are seen to be similar whether maturation occurs in vivo or in vitro. Capacitated sperm (both acrosome-intact and acrosome-reacted) are observed to interact with the microvilli of the oocyte surface. Little is known about oocyte surface proteins that mediate fertilization in mammals. Data of ours and others show that enzyme treatment of live unfertilized eggs interferes with sperm binding. Enzyme treatment (trypsin, chymotrypsin treatment, or pronase) reduces the number of bound sperm, suggesting removal of a surface protein involved in fertilization. Trypsin treatment also causes some lengthening of surface microvilli in a belt surrounding the metaphase II region. After metabolic labeling, proteins of zona-free unfertilized eggs can be identified by SDS-PAGE and autoradiography. Comparison of 1-D gels from untreated and enzyme-treated eggs show the nearly complete disappearance of proteins of 263, 170, 137, 97, and 87 kD after digestion; an increase in a 66 kD protein after trypsin or chymotrypsin; and a major new band of 20 kD after chymotrypsin treatment. Fertilized eggs show the loss of a 255-265 kD band among other changes. Proteins of 97 kD and 87 kD were seen previously by surface labeling (Johnson and Calarco, 1980b), and our 97 kD and 66 kD bands are similar in molecular weight to those identified by Boldt et al. (1989). Taken together, these data identify a few candidate proteins for the role of sperm receptor on the egg surface. Future work should focus on identification of the surface protein(s) which functions physiologically in fertilization by developing fertilization-blocking antibodies. Relatedness to other mammalian sperm receptors and identification of the genes involved would provide valuable information to our understanding of fertilization and to the problems of infertility and contraception.
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PMID:Fertilization of the mouse oocyte. 186 39

Antibody-mediated spermagglutination is responsible for infertility in some couples and fertilization in vitro can also be impaired by these antibodies. Having previously demonstrated the possibility of enzymatic disagglutination in such situations, the functional potential of disagglutinated spermatozoa has now been assessed. Chymotrypsin (500 U/ml) and papain (50 U/ml) resulted in impairment of oocyte penetration in the zona-free hamster egg penetration test. Trypsin (500 U/ml), while having no effect on egg penetration of normal spermatozoa, significantly improved oocyte penetration of spermatozoa which had been previously incubated with spermagglutinating antibody-positive sera. Sperm-mucus interaction was not improved, however, by trypsin treatment of agglutinated spermatozoa. This technique may be of value in conjunction with in-vitro fertilization in situations where spermagglutination exists, and also possibly with intra-uterine insemination if improved fertilizing ability can be confirmed in vitro.
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PMID:Treatment of spermagglutination with proteolytic enzymes. II. Sperm function after enzymatic disagglutination. 218 63

Alpha 2-macroglobulin (alpha 2M) is a plasma protein with proteinase inhibitor and immune modulatory capabilities. The amounts of alpha 2M in peritoneal fluid (PF) from women with endometriosis and women with noninflammatory gynecologic conditions were analyzed by functional (trypsin binding) and immunologic assays. The most important finding of this study was that a significant amount of the alpha 2M in the peritoneal fluid of patients with endometriosis had been inactivated by an as yet undetermined mechanism. The concentration and total amount of immunologically reactive alpha 2M in the samples varied widely and was not significantly different between the groups. However, women with endometriosis had significantly lower amounts of functional alpha 2M than did women without endometriosis. There was no significant difference between functional and immunologic measurements of alpha 2M in samples from women without endometriosis. Women with endometriosis, however, had less functional than immunologically reactive alpha 2M. This discrepancy was not due to inactivation of alpha 2M by having previously bound proteinase. Alpha 2M-proteinase complexes can down-regulate macrophase functions. It is possible that decreased proteinase-binding ability of alpha 2M may play a role in the pathogenesis of endometriosis and associated infertility by decreasing negative feedback control of macrophage activities.
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PMID:Reduced trypsin-binding capacity of alpha 2-macroglobulin in the peritoneal fluid of women with endometriosis: possible relevance to alterations in macrophase function. 245 48

After a brief review of the molecular structure of cervical mucus, the data are presented on inhibition of sperm transport through cervical mucus by polyanions and on enhancement of sperm penetration in cases of infertility due to antisperm antibodies. Cervical mucus is a gel made up of large, unbranched, glycosylated glycoprotein with highly glycosylated domains separated by hydrophobic peptide chains. Spermatozoa probably traverse the unbound water phase rather than the water bound to the macromolecules. Since mucin is a polyanion, polycations were investigated as potential vaginal spermicides. The two biguanides studies, chlorhexidine and Vantocil were both spermicidal in concentrations of 1-10 mg/ml. Their rate of entry into mucin in capillary tubes differed. Vantocil penetrated superficially and set up a barrier of inspissated mucus. Chlorhexidine entered further, with dept inversely proportional to concentration. Both biguanides increased the thickness of cervical mucus in a dose-dependent manner, as judged by dynamic storage modules, by sedimentation in analytical ultracentrifugation, and by solubility in 0.22 M thiocyanate. It was speculated that these biguanides act by altering the molecular configuration of mucin. In the presence of anti-sperm antibodies, spermatozoa observed in cervical mucus in vitro may show non-progressive mobility or immobility. The presence of auto-antibodies can be shown with Immunobeads. Binding of secretory IgA to sperm can be cleaved with bacterial protease as can binding of IgG with trypsin. By assaying the blockage of sperm by antibodies with Immunobeads and measuring penetration of sperm in donor cervical mucus, displacement of sperm antibodies could be demonstrated in 9 infertile subjects. Therefore, it might be possible to treat the ejaculate with proteases, and achieve conception by either a gamete intrafallopian tube transfer or an in vitro fertilization procedure.
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PMID:Changes in cervical mucus that prevent penetration by spermatozoa. 270 82

A sperm antigen has been isolated from radiolabeled human sperm cell membrane by detergent solubilization, lectin affinity chromatography, gel filtration, and indirect immune precipitation using sperm-immobilizing antisera from patients with unexplained infertility. Isolated material was characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Among 20 infertile women's sera with sperm-immobilizing antibodies, two were found to react predominantly with a sperm membrane polypeptide having the approximate molecular weight of 15,000 daltons. No significant binding to this molecule was observed in any sera from pregnant women, unmarried women, and normal men. By the absorption with spermatozoa, the antisera lost their binding activity to the molecule, while the sera absorbed with seminal plasma did not lose the activity. The results indicated that the molecule is a genuine sperm antigen and not a sperm-coating seminal plasma antigen. By the indirect immunofluorescence of washed ejaculated spermatozoa with the antisera, strong fluorescence was localized only in an equatorial segment of the acrosome, while no specific staining was observed in the controls. The antigen is relatively unstable against acid, alkali, and heat treatment. Treatment with proteolytic enzymes such as pronase and trypsin inactivated the antigen activity, indicating that the antigen epitope could be a peptide portion of the glycoprotein.
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PMID:Identification and characterization of a human sperm antigen corresponding to sperm-immobilizing antibodies. 320 36

This study determined the effects of Trichomonas vaginalis trophozoites, subcellular fractions, and medium from axenic T. vaginalis cultures on human sperm motility and viability. Spent medium (pH 7.0) caused complete cessation of sperm motility after 15 minutes incubation. Trophozoite soluble fraction or formalin-killed trophozoites caused a 50 percent reduction in sperm motility, compared to 25 percent reduction caused by the trophozoite particulate fraction or the sterile medium and three percent by saline (control). Spent medium from T. vaginalis cultures reaching stationary growth phase produced the greatest reduction in sperm motility, suggesting that potency was related to time in culture and trophozoites per ml. The T. vaginalis spermicidal activity was heat-stable, trypsin-sensitive, and had a molecular weight of 12-15,000 by gel filtration. This proteinaceous substance was present in and secreted by T. vaginalis trophozoites during normal growth in axenic culture. Since this T. vaginalis byproduct rapidly killed sperm in vitro, its effects in humans may contribute to infertility in infected couples.
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PMID:Trichomonas vaginalis: preliminary characterization of a sperm motility inhibiting factor. 326 54

Fractionation of human seminal plasma by high-speed centrifugation and by gel filtration on Sephadex G-100 produces a high molecular weight component which displays immunosuppressive characteristics. Both materials inhibit mitogen-induced blast transformation of normal human peripheral monocytes, the component isolated by gel filtration being more effective in this regard. Prior treatment with trypsin of the inhibitory material isolated by centrifugation virtually abolishes its inhibitory potency suggesting that an intact protein is essential for biological function. The results also suggest that suppression of the mitogen-induced blast transformation response of cultured monocytes, T- and B-cells can be reversed by the addition of freshly isolated T- or B-cells. The relevance of these experimental observations to problems of human infertility is discussed.
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PMID:In vitro studies in reproductive immunology: 4. Mechanisms of immune response control in human male genital tract. 614 9

Efforts were made to disrupt and solubilize human sperm cells and to evaluate the product for its content of infertility antibody-related antigens. In the procedure that was developed, a well-washed sperm sample is treated with 0.1 M dithiothreitol for 60 min, followed by trypsin at 0.1 mg/ml for 30 min, and then by soybean trypsin inhibitor. A mixture of DNAses I and II are added. After centrifuging, the resuspended pellet (RP) and the final supernatant (FS) show several degrees of cellular breakdown. Immunological evaluation was done with a strongly positive human serum containing sperm-head antibody. From the inhibition of sperm agglutination, we could conclude that the desired soluble antigen was obtained in the FS fraction.
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PMID:Human soluble antigens and their use in sperm antibody testing. 617 33

Human seminal plasma (SP) contains potent complement inhibitors. This study examined the complement-inhibiting activity of individual SP samples from 118 patients with infertility and analyzed them in relation to various semen parameters. When 25% complement-inhibiting activity was considered the cut off value, less than 1 SD unit from the mean percentage of inhibition of SP samples with normal semen quality, 32 samples (27%) showed low inhibiting activity. Among the lower group, incidences of patients with asthenozoospermia (66%) and oligozoospermia (31%) were significantly (p < .01) higher than those (36 and 10%) in the group whose SP showed significant inhibiting activity. Partial characterization revealed that the component responsible for complement inhibition was heat labile, trypsin resistant, high molecular weight (>10 kD) glycoprotein that can inhibit alternative as well as classical complement pathways. Furthermore, since in the majority of SP samples the anticomplementary activity was blocked by monoclonal antibody against membrane cofactor protein (MCP) or decay accelerating factor (DAF), the complement-inhibiting factors that were identified are likely to be MCP and/or DAF, which are known to be present in human SP. These results suggest that complement-regulatory proteins in SP such as MCP and DAF may protect sperm cells against complement attack in the male reproductive tract.
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PMID:Complement-inhibiting activity of human seminal plasma and semen quality. 890 71

In order to analyze the protective role that IgA may play in a chlamydial infection two IgA monoclonal antibodies (mAb), MoPn 4-2 and MoPn 13-2, were raised against the major outer membrane protein (MOMP) of the Chlamydia trachomatis mouse pneumonitis (MoPn) biovar. mAb MoPn 4-2 was found to be serovar specific while mAb MoPn 13-2 was species specific. mAb MoPn 4-2 recognized a surface exposed conformational epitope as shown by its ability to bind to native EBs and nonreduced MOMP while failing to bind to heat and trypsin treated EBs, to reduced MOMP and to synthetic MOMP peptides. In contrast, mAb MoPn 13-2 recognized a nonconformational epitope since it was able to bind treated EBs, to reduced MOMP and to the synthetic peptide MTTWNPTISGSGI located in variable domain 4 of the MOMP. Both mAbs agglutinated intact EBs and had in vitro neutralizing activity. However, mAb MoPn 4-2 had a 20-fold higher in vitro neutralizing ability when compared to mAb MoPn 13-2 (50% neutralization at 5 micrograms ml-1 vs 100 micrograms ml-1). In an in vitro in vivo infectivity assay, mAb MoPn 4-2 protected mice against infertility when C. trachomatis MoPn elementary bodies were preincubated with the mAb before inoculation. In addition, passive transfer of mAb MoPn 4-2 resulted in significant protection as measured by a decrease in the number of mice infected, and in the intensity and duration of vaginal shedding. These results support previous findings suggesting that IgA antibodies can play a role in protection against a chlamydial infection, and further encourage work to develop vaccination strategies that elicit mucosal immunity.
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PMID:Monoclonal immunoglobulin A antibody to the major outer membrane protein of the Chlamydia trachomatis mouse pneumonitis biovar protects mice against a chlamydial genital challenge. 916 May 28


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